Composition comprising melatonin or its derivatives with coenzyme q10 and use thereof against ageing of the skin

ABSTRACT

The invention relates to a composition comprising melatonin, a metabolite or derivative thereof and coenzyme Q10 and to use thereof for the production of a pharmaceutical or cosmetic composition for the treatment of the skin, said composition potentiating the movement of both molecules into the mitochondrion and facilitating percutaneous absorption, in which both the melatonin and the CoQ10 can reach all of the strata of the skin.

The present invention relates to use of a composition comprisingmelatonin, a derivative or a metabolite thereof, and coenzyme Q10suitable for potentiating the movement of said molecules into the cellmitochondrion and protecting the skin against the damage caused byoxidative stress. Therefore, the invention could be comprised in thefield of the pharmaceutical and cosmetic industry.

PRIOR STATE OF THE ART

The increase in the production of free radicals causes an increase inthe stiffness of cell membranes and connective tissue due to crosslinking. This leads to a decrease in blood supply to organs and tissues,with the subsequent decrease in the tissue perfusion. Free radicalsinexorably damage the cell and mitochondrial function slowly decreases,not only due to the oxygen radicals (ROS) that are generated within theactual organelle, but also due to those it receives from the cytosol. Asa result, the electron transport chain (ETC) and oxidativephosphorylation become inefficient, with the production of ATPdecreasing. This energy deficiency reduces the mitochondrial defensecapacity, which becomes more vulnerable to the attack of ROS, furtherdecreasing the efficiency thereof. A vicious circle is thus closed whichends with cell apoptosis.

In addition, ionizing radiations, such as ultraviolet radiation (UVR),are clear examples of ROS inducers, responsible for oxidative damage andimmune lesion of peripheral tissues such as the skin.

Melatonin is a neurohormone that is synthesized and metabolized in theskin, where it plays a very important role. Some of its functions arerelated to the antioxidant properties of melatonin and includeprotection against UV rays and X-rays, as well as to other oxidizingagents inducing oxidative damage in the skin, including surgicalinterventions and burns (Izykowska I et al., In Vivo 2009, 23:739-745).Particularly, melatonin is a strong protector against UV radiations,fundamentally against UVB radiations, which are the most noxiouscomponent of UV radiations. Exposure to both UV-B and UV-A inducesmelatonin metabolism with the subsequent increase in 2-hydroxymelatoninand in N1-acetyl-N2-formyl-5-methoxykynureneamine (AFMK). Furthermore,melatonin also activates the antioxidant cascade in the skin, reducingfree radicals. This data suggests that melatonin can control the redoxstate through its antioxidant functions.

The coenzyme Q10 (CoQ10; ubiquinone) is a molecule that is fundamentalfor mitochondrial function. It acts like an electron carrier in theelectron transport chain. The reduced form of CoQ10, or ubiquinol, isone of the most potent lipophilic antioxidants of cell membranes(Bentinger M et al., Mitochondrion 2007, 7, S41-S50). Furthermore, CoQ10is necessary for pyrimidine synthesis and can modulate apoptosis andmitochondrial uncoupling proteins.

CoQ10 increases mitochondrial function and is therefore useful inmitochondrial pathologies and in pathologies presenting withmitochondrial damage, such as neurodegenerative diseases (Parkinson'sand Alzheimer's), as well as in cellular ageing.

Skin cells depend on mitochondrial activity to produce enough energy forcell proliferation, mitochondrial DNA replication, protein synthesis andother cellular activities to repair the damage caused by theenvironment. Mitochondrial function and ATP production in fibroblastsdrop with exposure to UV radiations (Berneburg M et al., J InvestDermatol 2005, 125:213-220). In other studies it has been demonstratedthat levels of CoQ10 in the skin decrease with age. Therefore, thetopical application of Co10 could have very beneficial effects for theskin.

A number of papers have evaluated the efficacy of the administration ofCoQ10 to normalize mitochondrial function (López L C et al., Plos One2012, doi:10.1371/journal.pone.0011897); however, CoQ10 is a verylipophilic molecule that accumulates in the membrane and moves into themitochondrion in very small amounts. Therefore, exogenous CoQ10 isprimarily distributed in the lysosomes, endoplasmic reticulum and plasmamembrane, and only a small amount moves into the mitochondrion.Furthermore, most of the CoQ10 that moves into the mitochondrion istrapped by the external mitochondrial membrane, not being available forthe respiratory chain which is located in the internal mitochondrialmembrane. The administration of CoQ10 does not increase levels of ATP inpatients with a CoQ10 deficiency after one week of treatment.

Compositions for the treatment of ageing of the skin, comprising themixture of several substances, and included among them are bothmelatonin and coenzyme Q10, have been claimed in various patents.

US patent application 2005/0025756 A1 describes the combination of CoQ10and other chemical substances in a chemical composition causing anincrease in the delivery of coenzyme Q10 into a cell, which enablesovercoming the solubility problems thereof, being able to be absorbed bythe cell more quickly.

Accordingly, there is a need for a tool that facilitates the movement ofCoQ10 into the mitochondrion and potentiates the movement of melatonininto the mitochondrion, and thereby reverses the mitochondrial damagecaused in cellular ageing as well as in many pathologies presenting withmitochondrial damage.

DETAILED DESCRIPTION OF THE INVENTION

The present invention describes a composition comprising melatonin or aderivative thereof, preferably at a concentration of 1.0 to 5.0% w/v,and CoQ10, preferably at a concentration of 0.2 to 1.5% w/v, for theproduction of a pharmaceutical or cosmetic composition for potentiatingthe movement of said molecules into the mitochondrion, and therefore usethereof for the treatment and/or prevention of ageing of the skin.

The examples of the following invention show in vivo results of thelevels of these molecules in the cytosol and in the mitochondria of skincells, as well as the plasma levels reached after application of thecream on the skin. The composition of the invention is useful forpotentiating the movement of CoQ10 and melatonin into the mitochondrionand for favoring percutaneous absorption, assuring that the melatoninand CoQ10 reach all of the layers of the skin, epidermis and dermis. Thepresent invention demonstrates that the combination of CoQ10 andmelatonin increases the movement of both molecules into the cell, andprimarily into the mitochondrion, allowing percutaneous absorption atthe same time, which does not occur when creams with the individualcompounds are applied.

The usefulness of the combination of melatonin, or its derivatives, andCoQ10 in the topical application on the skin, with a cell protectioneffect against oxidative stress, can be inferred from this earlier data.The topical application of melatonin+CoQ10 prevents cell functionimpairment as a result of mitochondrial damage caused by oxidativestress.

Therefore, a first aspect of the present invention relates to acomposition (hereinafter “composition of the invention”) comprising:

a. coenzyme Q10 (CoQ10), and

b. at least one compound of general formula (I),

where:n is an integer between 1 and 4; preferably 1, 2 or 3.R₁ is a straight or branched (C₁-C₄) alkyl group;R₂ is hydrogen, straight or branched (C₁-C₄) alkyl, a —C(═O)O—Ra groupor a —C(═O)—N(H)—Ra group, wherein Ra is a straight or branched (C₁-C₄)alkyl group; andR₃ is a hydrogen or a straight or branched (C₁-C₄) alkyl group.

The expression “melatonin, a metabolite or derivative thereof” isunderstood as any cosmetically or pharmaceutically acceptable compoundthat is comprised within general formula (I), as well as the salts,solvates or prodrugs thereof, and is useful for the production of apharmaceutical or cosmetic composition with CoQ10 for potentiating themovement of this molecule into the mitochondrion and facilitatingpercutaneous absorption. Therefore, when the present invention refers tothe compounds of general formula (I), the present invention also refersto the salts, solvates or prodrugs thereof. Therefore, the compounds ofgeneral formula (I) described above refer to melatonin, the metabolitesor derivatives thereof.

The term “alkyl” herein refers to aliphatic, straight or branchedchains, having from 1 to 4 carbon atoms, for example, methyl, ethyl,n-propyl, i-propyl, n-butyl, tert-butyl, sec-butyl, etc. Preferably thealkyl group has between 1 and 2 carbon atoms, and more preferably it isa methyl group.

In a preferred embodiment of the present invention R₁ is a methyl groupand R₃ is hydrogen or a methyl group.

In another preferred embodiment n is 1.

Preferably in another embodiment R₂ is hydrogen.

In another preferred embodiment the compound of formula (I) is selectedfrom melatonin or N-acetylserotonin. More preferably it is melatonin.

The term “melatonin” refers to N-acetyl-5-methoxytryptamine, alsoreferred to in the literature as melatonin, melatonine, melatol,melovine, circadin, regulin, acetaminde, N-acetyl-methoxy-tryptamine,5-methoxy-N-acetyltryptamine,N-[2-(5-methoxy-1H-indol-3-yl)ethyl]acetamide orN-[2-(5-methoxyindol-3-yl)ethyl]acetamide, or when in the compound ofgeneral formula (I) R₁ and R₃ are a methyl group, n is 1 and R₂ ishydrogen. The CAS number for melatonin is the 73-31-4.

Melatonin is an endogenous neurohormone produced by the pineal gland(epiphysis cerebri), as well as by other organs and tissues, such as thegastrointestinal tract, the retina, lymphocytes and cells of the bonemarrow, and the skin, for example, in a physiological manner in animals,inter alia, in humans.

Melatonin is produced in animals, inter alia, humans, from serotonin(5-hydroxytryptamine, 5-HT), which in turn derives from the amino acidtryptophan. Therefore, the present invention could also relate to use ofa composition comprising any of the melatonin precursors (5-HT,tryptophan or intermediate metabolites such as N-acetylserotonin, orNAS), at a sufficient concentration so that in the human body they aretransformed into melatonin at the concentrations described in thepresent invention, for the production of a pharmaceutical compositionwith CoQ10 for potentiating the movement of this molecule into themitochondrion and facilitating percutaneous absorption.

In addition, the present invention also relates to the pharmaceuticallyacceptable salts of melatonin or of its derivatives which can begenerated by means of chemical processes known by the person skilled inthe art, for example, by means of a reaction with an acid in water or inan organic solvent or in a mixture of both. Ether, ethyl acetate,ethanol, isopropanol or acetonitrile can be used as an organic solvent.Examples of acid addition salts include mineral acid addition salts suchas, for example, hydrochloride, hydrobromide, hydroiodide, sulfate,nitrate, phosphate, and organic acid addition salts such as, forexample, acetate, maleate, fumarate, citrate, oxalate, succinate,tartrate, malate, mandelate, methanesulfonate and p-toluenesulfonate.

The term “prodrug”, as it is used herein, relates to a chemical compoundthat has experienced a chemical derivation, for example, a substitutionor an addition of an additional chemical group, to modify any of itsphysicochemical properties, such as solubility or bioavailability, butwhich does not modify the technical characteristics of the originalmolecule. A prodrug may be, for example, an ester, ether or amidederivative. Bioavailability relates to the availability thereof in aspecific biological compartment.

The term “solvate” according to this invention must be understood asthat derivative of melatonin that has another molecule, for example, apolar solvent, attached by means of a non-covalent bond. The examples ofsuch solvates include hydrates and alcoholates, for example,methanolates.

The preparation of solvates and prodrugs can be carried out by means ofmethods known in the state of the art.

Non-pharmaceutically acceptable salts, solvates or prodrugs are alsowithin the scope of the invention since they can be useful in thepreparation of pharmaceutically acceptable salts, solvates or prodrugs.

The composition of the invention can also relate to a compositioncomprising a functional biological equivalent of melatonin at aconcentration which is equivalent to that described in the compositionsof the invention.

The term “functional biological equivalent” or “bioequivalent variable”,as it is used herein, refers to a molecule with the same function as thedescribed molecule, which can have slight variations with respect to thedescribed molecule without said variations providing any added technicaleffect for said molecule. The present invention therefore refers tomelatonin variants having the same function and having slight variationswithout said variations providing any added technical effect formelatonin.

“Concentration that is equivalent” is understood as that concentrationnecessary for the functional biological equivalent of melatonin toproduce the same effect as that described in the present invention forthe composition of the invention.

Melatonin is also produced in plants. For example, the presence ofmelatonin has been described in algae, edible plants, cereals, fruits,seeds, roots, stems, leaves and medicinal herbs. For example, thepresence of melatonin has been described in cocoa, coffee, grapes,tomatoes, green tea, algae, cereals and olives, so the origin of themelatonin of the composition of the invention can be from a plantorigin. Obtaining the melatonin from a plant origin (also referred to asphytomelatonin) can be done by means of any method known by the personskilled in the art for such purpose.

The origin of the melatonin used in the composition of the invention canalso be synthetic. Chemical synthesis of melatonin can be done by meansof the techniques known by the person skilled in the art for suchpurpose.

The most common form of CoQ in humans is CoQ10, with the followingstructural formula:

although small traces of CoQ9 can also be found. This molecule is alsoreferred to as ubiquinone since it is produced by virtually allorganisms with respiratory metabolism. It is a fat-soluble benzoquinone.The “Q” refers to the quinone chemical group, and the “10” refers to thenumber of isoprenoid subunits it has. The benzoquinone portion of CoQ10is synthesized from tyrosine, whereas the isoprenoid chain issynthesized from acetyl-CoA through the mevalonate pathway. The varioustypes of coenzyme Q can be distinguished by the number of isoprenoidchains they have.

In another preferred embodiment, the composition of the inventioncomprises a compound of formula (I), preferably melatonin, at aconcentration of 1 to 5% w/v with respect to the final composition, morepreferably at a concentration of 1.5 to 3% w/v with respect to the finalcomposition, and even more preferably at a concentration of about 2% w/vwith respect to the final composition.

In another preferred embodiment, the composition of the inventioncomprises CoQ10 at a concentration of 0.2 to 1.5% w/v with respect tothe final composition, more preferably at a concentration of 0.3 to 1.0%w/v with respect to the final composition, and even more preferably at aconcentration of about 0.5% w/v with respect to the final composition.

In another preferred embodiment, the composition of the inventioncomprises a compound of formula (I), preferably melatonin, at aconcentration of 1.0 to 5.0% w/v, plus CoQ10 at a concentration of 0.2to 1.5% w/v.

“Concentration of 1 to 5% w/v” is understood as the compositioncomprising from 1 to 5 grams of the compound of formula (I), preferablymelatonin, in 100 ml of final composition.

“Concentration of 0.2 to 1.5% w/v” is understood as the compositioncomprising from 0.2 to 1.5 grams of CoQ10 in 100 ml of final composition

The abbreviation “w/v” refers to weight/volume or mass/volume (m/v).

In a particular embodiment, the composition of the invention comprisesat least one antioxidant.

The term “antioxidant” refers to that substance that is able to delay orprevent oxidation, particularly of the compound of general formula (I)and more particularly of melatonin. Antioxidizing agents known in thestate of the art can be used as antioxidizing agents, for example, butwithout being limited, the list comprising tocopherol, ascorbic acid,sodium ascorbate, tartaric acid, butylhydroxyanisole, citric acid,vitamin A and vitamin E.

In a more particular embodiment of the preceding one, the antioxidant isat a concentration of 0.25 to 0.75% w/v with respect to the finalcomposition. More preferably the concentration is from 0.40 to 0.60% w/vwith respect to the final composition.

In another particular embodiment of the composition of the invention itfurthermore comprises at least one preservative.

“Preservative” is understood as that substance maintaining theproperties of the cosmetic or medicinal product by inhibitingcontamination by germs; it can be an ionic or non-ionic preservative.The preservative used will not be toxic, will be chemically stable andcompatible with both the compound of formula (I), particularlymelatonin, and with CoQ10. The preservative is preferably at aconcentration of 0.4 to 1% w/v with respect to the final composition.Preservative agents known in the state of the art can be used aspreservative agents, for example, but without being limited, thepreservative can refer to benzoic acid, sodium benzoate, ascorbic acid,potassium sorbate, methylparaben, ethylparaben or butylparaben. “Germs”are understood as any cell that can grow and multiply in the compositionof the invention, for example, bacteria, fungi and yeasts.

Another particular embodiment of the composition of the inventionfurthermore comprises at least one gelling agent. The gelling agent ispreferably at a concentration of 2.5 to 7% w/v with respect to the finalcomposition.

The term “gelling agent” refers to a substance that forms a gel, i.e., athree-dimensional network formed by the gelling agent and generallycontaining a liquid phase. The gelling agents that can be used can bethose known by the person skilled in the art for the production of apharmaceutical or cosmetic composition. Preferably the gelling agent isselected from the list comprising polyethylene and polypropylenecopolymer, cellulose and guar gum.

In a particular embodiment, the composition of the invention furthermorecomprises at least one “pharmaceutically and/or cosmetically acceptablevehicle”, which refers to those substances, or combination ofsubstances, known in the pharmaceutical and/or cosmetic sector used inthe production of cosmetic and/or pharmaceutical forms of administrationand includes, but without being limited, solids, liquids, solvents orsurfactants. The vehicle can be an inert substance or a substance havingan action that is analogous to any of the compounds of the presentinvention. The vehicle is preferably at a concentration of 2.5 to 7% w/vwith respect to the final composition. The function of the vehicle is tofacilitate incorporation of the expression product of the invention, aswell as also other compounds, to allow better dosing and administrationor to provide consistency and form to the pharmaceutical composition.When the presentation form is liquid, the vehicle is the diluent. Thesevehicles can be known by the person skilled in the art, for example,lysosomes, millicapsules, microcapsules, nanocapsules, sponges,millispheres, microspheres, nanospheres, milliparticles, microparticlesand nanoparticles.

In a particular embodiment, the composition of the invention furthercomprises at least one pharmaceutically and/or cosmetically stableexcipient or adjuvant. The excipient is preferably at a concentration of1 to 5% w/v with respect to the final composition.

The term “excipient” refers to a substance which aids in the absorptionof the pharmaceutical or cosmetic composition of the invention,stabilizes said compositions or aids in its preparation in the sense ofgiving it consistency or providing flavors making it more pleasant.Therefore, the excipients may have the function of keeping theingredients bound to one another, such as starches, sugars orcelluloses, for example, a sweetening function, a colorant function, amedicinal product protection function, such as insulating it from theair and/or moisture, for example, a filler function of a pill, capsuleor any other presentation form, such as dibasic calcium phosphate, forexample, a disintegrating function for facilitating dissolution of thecomponents and absorption thereof into the intestine, without excludinganother type of excipients not mentioned in this paragraph, for examplestarches, sugars or celluloses. In order for the composition of theinvention to have a pleasant flavor, an essence, such as essence ofcinnamon, lemon, orange, mandarin or vanilla, for example, can be added.

The term “adjuvant” refers to any substance potentiating the response ofan active ingredient. In the present invention it refers to anysubstance potentiating the effects of the composition of the invention;it can refer to any adjuvant known by the person skilled in the art.

The term “pharmaceutically and/or cosmetically acceptable” refers to thecompound to which reference is made being allowed and evaluated suchthat it does not cause damage to the organisms to which it isadministered.

In another particular embodiment, the composition of the inventionfurthermore comprises at least other additives such as an emulsifier, anemollient or an antifoaming agent. In the composition of the inventioneach of these additives can individually be at a concentration of 2.5 to7% w/v with respect to the final composition. Therefore, said particularcomposition refers to a composition comprising between 2.5 and 7.0 gramsof additive in 100 ml of total volume of the final composition.

“Emulsifier” is understood as those substances the function of which isto stabilize mixtures of two immiscible liquids. The list of emulsifiersof the particular embodiment includes Montanov 68, glycerol, sodiumlauroyl glutamate, sodium cocoyl glutamate, PEG-26 jojoba acid, PEG-26jojoba alcohol, PEG-11 avocado glycerides, PEG-30 almond glycerides,caprylic compound and triethylhexanoin.

“Emollient” is understood as a complex mixture of chemical agentsparticularly designed to act at different levels in epidermal strata,for softening and slackening tissue. The action thereof can besuperficial or deep, according to if the composition remains on thesurface of the skin or if it is absorbed. The emollient for theparticular embodiment can be selected from the list comprisingperhydrosqualene, lanolin, petroleum jelly, rosehip oil, borage oil,sweet almond oil, peach kernel oil, olive oil, beeswax, paraffin waxes,stearyl alcohol and cetyl alcohol.

“Antifoaming agent” is understood as an additive used in the cuttingfluids to aid in preventing foaming and bubbling, and to thus improvethe cooling capacity of the fluid. They are products with low viscosityand ease for being quickly propagated on surfaces with effervescencewhere they destabilize the foam films, which causes the air bubbles toburst and the breakdown of the foam of the surface. The antifoamingagent of this particular embodiment is selected from the list comprisingdimethicone, dimethiconol, phenethyl disiloxane, tetramethyl decynediol,phenyl trimethicone, polysilicone 7, isopropyl alcohol, hexyl alcohol,propyl alcohol, hexamethyl-disiloxane, bisphenyl hexamethicone andtrimethyl-siloxy silicate.

In another particular embodiment, the composition further comprises atleast one antimicrobial agent.

The term “antimicrobial agent” describes a substance acting againstparasitic microorganisms such as bacteria, viruses, or fungi, by killingthem or inhibiting their growth. The antimicrobial agent is selectedfrom the list comprising phenonip (composition the main component ofwhich is 2-phenoxyethanol), methylparaben, ethylparaben, butylparaben,propylparaben, methyl parahydroxybenzoate, propyl parahydroxybenzoate,benzalkonium chloride and imidazolidinyl-urea.

In an even more particular embodiment of the preceding embodiment, theantimicrobial agent is at a concentration of 0.4 to 1.0% w/v withrespect to the final composition. In other words, said particularcomposition refers to the composition of the invention furthercomprising at least between 0.4 and 1.0 grams of additive in 100 ml oftotal volume of the final composition.

In another particular embodiment, the composition of the inventionfurther comprises at least one agent acting as a filter of UV rays,preferably UV-B and/or UV-A rays. This is understood as those substanceswhich aid in preventing sunburns and reduce the harmful effects of thesun, such as premature ageing of the skin and skin cancer. Among thosemost widely used, stand out zinc oxide, titanium dioxide, iron oxide,talc, silicates (clays, kaolin, bentonite), PABA (p-aminobenzoic acid)and derivatives, cinnamates, anthranilates, salicylates, benzophenones,dibenzoylmethane and benyzlidene-camphor.

The composition of the invention can be both a cosmetic composition anda pharmaceutical composition, and it can be formulated foradministration in a range of forms known in the state of the art. Suchformulations can be administered to an animal, and preferably to amammal, and more preferably to a human, through a range of routes,including, but without being limited to topical route, oral route,parenteral route, intraperitoneal route, intravenous route, intradermalroute, intralesional route, intraarterial route, intramuscular route,intranasal route, or subcutaneous route.

Based on what is herein described, another particular embodiment relatesto use where the composition is presented in a form adapted for topicaladministration.

The term “topical administration” herein refers to the composition beadministered on the skin.

A particular embodiment of the composition of the invention relates touse thereof in the form of a cream, lotion, gel or aerosol. In additionto the preceding formulations, the following can also be used in thecomposition of the invention: oil in water emulsions, water in oilemulsions, milks, lotions, gels, ointments, balsams, foams, body oils,soaps, bars, pencils, vaporizers, creams, liniments, salves, sera andmousses. The composition can also be incorporated in solid supportsselected from the group consisting of hydrogels, wipes, patches andfacial masks.

In another particular embodiment, the composition of the invention is inthe form of an emulsion. “Emulsion” is understood as a more or lesshomogenous mixture of immiscible liquids. One liquid (the dispersedphase) is dispersed in another one (the continuous phase or dispersingphase). The emulsion of the invention is an O/W (oil/water) emulsionwhich is described as the dispersed phase being an oily phase and thedispersing phase being an aqueous phase.

In another particular embodiment, the composition of the inventioncorresponds with a pharmaceutical or cosmetic composition.

“Cosmetic composition” or “cosmetic” is understood as any substance ormixture intended for being placed in contact with superficial parts ofthe human body (epidermis, hair and capillary system, nails, lips, andexternal genital organs) or with the teeth and buccal mucosae, for theexclusive or primary purpose of cleaning them, perfuming them, changingtheir appearance, protecting them, keeping them in good condition orcorrecting body odors.

The term “pharmaceutical composition” or “medicinal product” refers toany substance used for the prevention, diagnosis, alleviation, treatmentor curing of diseases in humans or animals. In the context of thepresent invention it refers to a composition capable of treating and/orpreventing skin pathologies.

Another aspect of the invention relates to the use of the composition ofthe invention and of the compositions of the particular embodiments forthe production of a medicinal product.

A particular embodiment of the aspects and preceding particularembodiments, as well as of the composition described in each of theparticular embodiments, corresponds with the use thereof for thetreatment and/or prevention of cellular senescence or of diseasespresenting with cellular ageing. “Cellular senescence” is understood asthe process initiated in response to stress and damage occurring in acell, and it constitutes an alternative programmed cell death responsepathway and is of vital importance for suppressing cancer cellformation. It is also associated with the repair of tissues andinflammation thereof, processes associated with tumor growth. Thecellular senescence is therefore simultaneously associated with tumorsuppression and promotion processes, like in the ageing and repair oftissues.

In a particular embodiment, diseases presenting with cellular ageing maytherefore be tumors, cancers, inflammation of tissues, and moreparticularly those related to the skin.

In the present invention “treatment and/or prevention” refers to boththerapeutic treatment and prophylactic treatment; or preventivemeasures. Those situations susceptible to treatment include thosealready associated with disorders as well as in those in which thedisorder is prevented. A “disorder” is any condition that would benefitfrom treatment with the composition of the invention, as it is describedherein.

Another aspect of the invention relates to a cosmetic method foranti-ageing of the skin, which is characterized by the topicalapplication, preferably in the form of a cream or emulsion, of atherapeutically effective amount of the composition of the invention.

“Anti-ageing” herein refers to the process of slowing down, preventingor reversing the ageing process.

The expression “therapeutically effective amount” of the compositionrefers to the pharmaceutically or cosmetically effective amount of thecomposition that produces the desired effect, and it is generallydetermined, among other causes, by the characteristics typical of saidpharmaceutical composition and of the therapeutic effect to be achieved.The dosage for obtaining a therapeutically effective amount depends on arange of factors, such as, for example, age, weight, sex or tolerance ofthe animal, preferably a mammal and more preferably human.

“Ageing of the skin” is a process related to the loss of fibrous oftissue, the slowing down of the cell turnover rate and decrease inglandular and vascular activity in cutaneous tissue. Changes in skin dueto the ageing process are related to environmental factors, such asexposure to the sun, genetic constitution, nutrition, passage of time,inter alia. Changes in connective tissue reduce skin resistance andelasticity, causing unwanted effects in skin such as wrinkles, liverspots, among others known by any person skilled in the art.

In a particular embodiment of this aspect of the invention, ageing ofthe skin is caused by exposure of the skin to the sun's rays.

Therefore, another aspect of the present invention relates to a cosmeticcomposition comprising the composition of the invention, and moreparticularly said composition is a sunscreen, which preferably is acream, lotion, gel, milk or oil.

Throughout the description and claims the word “comprises” and itsvariants do not seek to exclude other technical characteristics,additives, components or steps. For the persons skilled in the art,other objects, advantages and features of the invention will be inferredin part from the description and in part from the practice of theinvention. The following examples and drawings are provided by way ofillustration and do not intend to be limiting of the present invention.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows levels of CoQ10 in cytosol of the skin of mice treated withcream comprising melatonin, cream comprising CoQ10, and cream comprisingmelatonin+CoQ10. Controls (C), mice treated with cream comprisingmelatonin (aMT), mice treated with cream comprising CoQ10 (CoQ), andmice treated with cream comprising melatonin+CoQ 10 (aMT+CoQ).***p<0.001 compared to C; +p<0.05 compared to CoQ10, ###P<0.001 comparedto melatonin.

FIG. 2 shows levels of CoQ10 in mitochondrion of the skin of micetreated with cream comprising melatonin, cream comprising CoQ10, andcream comprising melatonin+CoQ10. Controls (C), mice treated with creamcomprising melatonin (aMT), mice treated with cream comprising CoQ10(CoQ), and mice treated with cream comprising melatonin+CoQ 10(aMT+CoQ). ***p<0.001 compared to C; ++p<0.01 compared to CoQ10,###P<0.001 compared to melatonin.

FIG. 3 shows levels of CoQ10 in the plasma of mice treated with creamcomprising melatonin, cream comprising CoQ10, and cream comprisingmelatonin+CoQ10. Controls (C), mice treated with cream comprisingmelatonin (aMT), mice treated with cream comprising CoQ10 (CoQ), andmice treated with cream comprising melatonin+CoQ 10 (aMT+CoQ).***p<0.001 compared to C; +++p<0.001 compared to CoQ10, ###P<0.001compared to melatonin.

FIG. 4 shows levels of melatonin in cytosol of the skin of mice treatedwith cream comprising melatonin, cream comprising CoQ10, and creamcomprising melatonin+CoQ10. Controls (C), mice treated with creamcomprising melatonin (aMT), mice treated with cream comprising CoQ10(CoQ10), and mice treated with cream comprising melatonin+CoQ 10(aMT+CoQ 10). ***p<0.001 compared to C; +++p<0.001 compared to CoQ10,###P<0.001 compared to melatonin.

FIG. 5 shows levels of melatonin in mitochondrion of the skin of micetreated with cream comprising melatonin, cream comprising CoQ10, andcream comprising melatonin CoQ10. Controls (C), mice treated with creamcomprising melatonin (aMT), mice treated with cream comprising CoQ10(CoQ), and mice treated with cream comprising melatonin+CoQ 10(aMT+CoQ). ***p<0.001 compared to C; +++p<0.001 compared to CoQ10,###P<0.001 compared to melatonin.

FIG. 6 shows levels of melatonin in the plasma of mice treated withcream comprising melatonin, cream comprising CoQ10, with creamcomprising melatonin+CoQ10. Controls (C), mice treated with creamcomprising melatonin (aMT), mice treated with cream comprising CoQ10(CoQ), and mice treated with cream comprising melatonin+CoQ 10(aMT+CoQ). ***p<0.001 compared to C; +++p<0.001 compared to CoQ10,###P<0.001 compared to melatonin.

EMBODIMENTS

The following examples provided in this patent document serve toillustrate the nature of the present invention. These examples areincluded only for illustrative purposes and must not be interpreted aslimitations to the invention herein claimed. Therefore, the examplesdescribed below illustrate the invention without limiting the field ofapplication thereof.

The invention will be illustrated below by means of tests performed bythe inventors which clearly show the usefulness of the cream comprisingmelatonin plus CoQ10 for potentiating the movement of these compoundsinto cells, primarily into the mitochondrion, and favoring percutaneousabsorption, allowing these compounds to reach all of the layers of theskin.

Example 1

The following example provides formulations of cream comprisingmelatonin, cream comprising CoQ10 and cream comprising melatonintogether with CoQ10 and the method of producing them:

CREAM 1: MELATONIN (aMT) PHASE A: 70° C. Montanov 68 5% Perhydrosqualene5% Long-chain TG 5% Dimethicone 5% Phenopip 0.80%   Vitamin E 0.50%  PHASE B: 75° C. Glycerol 5% H₂O qsf Melatonin 2% in 30% PG CREAM 2:CoQ10 PHASE A: 70° C. Montanov 68 5% Perhydrosqualene 5% Long-chain TG5% Dimethicone 5% Phenopip 0.80%   Vitamin E 0.50%   PHASE B: 75° C.Glycerol 5% H₂O qsf CoQ10 0.5% in 5% PG The solution comprising CoQ isadded to the emulsion at 60° C. CREAM 3: CoQ10 + aMT PHASE A: 70° C.Montanov 68 5% Perhydrosqualene 5% Long-chain TG 5% Dimethicone 5%Phenopip 0.80%   Vitamin E 0.50%   PHASE B: 75° C. Glycerol 5% H₂O qsfMelatonin 2% in 30% PG CoQ10 0.5% in 5% PG The solution comprising CoQis added to the emulsion at 60° C.where: TG triglyceride; PG propylene glycol.

The method for producing the creams 1, 2 and 3 was carried out by meansof the following steps:

Cream 1:

PHASE A: all the components of phase A are mixed and it is heated untilreaching the temperature of 70° C.

PHASE B: the 2% melatonin in 30% PG is prepared: 2 grams of melatoninare weighed and dissolved in 30 ml of PG for 100 ml of finalcomposition. All the components are mixed and it is heated untilreaching the temperature of 75° C.

Finally, phase B is added to phase A.

For creams 2 and 3, the method is the same as for cream 1.

Phase B is added to phase A. The solution comprising CoQ in PG is addedwhen the temperature of this mixture reaches 60° C.

Example 2

Each of the creams described in the preceding example was administeredto experimentation animals by topical route. Nude male mice 3 months ofage (Harlan) were used. The animals were independently housed inconventional cages in an animal facility conditioned for that purpose inthe Centro de Investigación Biomédica (Biomedical Research Center), insterile conditions and under a controlled light period (12:12 h) andtemperature (22±1° C.) environment, and laboratory diet and water adlibitum. The study was conducted in samples of skin from the back of themice.

The experimental groups included 6 mice each, and they were thefollowing:

-   -   Control group (C)    -   Group treated with cream comprising 2% melatonin    -   Group treated with cream comprising 0.5% CoQ10    -   Group treated with cream comprising 2% melatonin+0.5% CoQ10

Application of the different formulations of the cream was done in thedorsal area at a ratio of 15 mg/cm²/day (5 mg/cm²/administration).

Administration of the cream in the different groups was done twice a day(every 12 hours). The animals were sacrificed a month after startingtreatment.

Blood samples are taken before the animals are sacrificed. The animalsare anesthetized with chloroform and 1 ml of blood is taken by means ofcardiac puncture. Plasma is separated from the red blood cells, which isused to measure levels of melatonin. After taking the blood sample andsacrificing the animals, samples of skin are taken from the back bymeans of surgical dissection. The skin is placed in a Petri dish withphosphate-buffered saline. The adipose tissue is removed, cut intopieces, and frozen at −80° C. until analysis. The method for extractingthe epidermis and the histochemical technique are based on the protocoldescribed by Scaletta and MacCalum, (Am J. Anat. 133: 431-53, 1972).

These samples were used to evaluate levels of melatonin and CoQ10 in thecytosol and mitochondrion of the skin and in plasma.

By observing the results obtained in FIGS. 2 and 5, it is shown thatmelatonin potentiates the movement of CoQ10 into the mitochondrion, andconversely, CoQ10 potentiates the movement of melatonin into saidorganelle, since both compounds are at a significantly higher amount inthe mitochondrion in the group in which they were jointly administeredin the cream, compared with the administration of only melatonin or ofonly CoQ10. FIG. 2 shows that melatonin potentiates the movement ofCoQ10 into the mitochondrion. FIG. 5 shows that CoQ10 potentiates themovement of melatonin into the cell in a very significant manner. Aconcentration of melatonin in the mitochondrion in the presence of CoQ10is shown to be 4 times greater. These results are confirmed by what isshown in FIGS. 1 and 4, since in the case of FIG. 1, levels of CoQ10 inthe cytosol are lower in the group of the joint administration ofmelatonin and CoQ10, than in the case of CoQ10 administered alone.Therefore, in the case of joint administration, CoQ10 began to beintroduced into the mitochondrion from the cytosol. In the case of FIG.4, CoQ10 potentiates in a very significant manner the movement ofmelatonin into the cell, and specifically into the mitochondrion.Additionally, despite the fact that CoQ10 increases the movement ofmelatonin into the mitochondrion, the melatonin continues to be veryhigh in the cytosol, levels of this molecule in the cytosol being muchhigher when it is administered together with CoQ10 than when it isadministered alone.

In addition to the movement of CoQ10 into the mitochondrion beingpotentiated, percutaneous absorption of this molecule is simultaneouslyfavored as melatonin and CoQ10 are administered at the same time. Thisis reflected in the results shown by the group of joint administrationin FIGS. 3 and 6, in which the levels of both CoQ10 and melatonin,respectively, are higher in plasma. Melatonin allows the absorption ofCoQ through the dermis, which means that this molecule can reach all ofthe layers of the skin and does not remain only in the epidermis.

1-55. (canceled)
 56. A composition comprising: a) coenzyme Q10 at aconcentration of 0.3 to 1.0% w/v with respect to the final composition,b) melatonin at a concentration of 1.0 to 5% w/v with respect to thefinal composition and c) at least one antioxidant at a concentration of0.25 to 0.75% w/v with respect to the final composition.
 57. Thecomposition according to claim 56, wherein the melatonin is at aconcentration of 1.5 to 3% w/v with respect to the final composition.58. The composition according to claim 56, wherein the antioxidant isselected from the group consisting of ascorbic acid, sodium ascorbate,tartaric acid, butylhydroxyanisole, citric acid, vitamin A and vitaminE.
 59. The composition according to claim 56 further comprising at leastone gelling agent.
 60. The composition according to claim 56 furthercomprising at least one pharmaceutically or cosmetically acceptablevehicle, and/or at least one pharmaceutically or cosmetically acceptableexcipient or adjuvant.
 61. A method for treating and preventing celldamage caused by oxidative stress in a patient in need thereof, saidmethod comprising topically administering to the patient the compositionaccording to claim
 56. 62. A method for treating and preventing adisease presenting cellular aging in a patient in need thereof, saidmethod comprising topically administering to the patient the compositionaccording to claim
 56. 63. The method according to claim 62, wherein thedisease presenting cellular aging is selected from the group consistingof tumors, cancers, inflammation of tissues and diseases related to theskin.
 64. The method according to claim 62, wherein cellular aging isdue to exposure to the sun in the skin.
 65. A cosmetic method foranti-aging of the skin of a subject, comprising topical application ofan effective amount of the composition according to claim 56 to the skinof the subject.
 66. The cosmetic method according to claim 65, whereinthe aging of the skin is caused by the exposure of the skin to the sun'srays.
 67. A sunscreen comprising the composition according to claim 56and at least one agent acting as filter for UV-B and UV-A rays.
 68. Thecomposition according to claim 56, wherein the antioxidant istocopherol.